Reduction of total hemocyte count (THC) in Cr(VI) exposed <i>D. melanogaster</i> larvae.

<p>Graphical representation of total hemocyte number (%) in Cr(VI) exposed Oregon R⁺ larvae after immunostaining by Hemese (H2) antibody (A). Hemocyte population in <i>hml^Δ-Gal4 UAS-2xEGFP</i> larvae after Cr(VI) exposure (B). Graph representing total hemocyte count in Cr(VI) exposed <i>hml^Δ-Gal4 UAS-2xEGFP</i> larvae as determined by flow cytometry (C). Data represent mean ± SD (n = 3) (20 larvae in each replicate). Significant differences were ascribed as *<i>p</i><0.05; **<i>p</i><0.01 and ***<i>p</i><0.001 as compared to control. Representative confocal microscopic images of the hemocytes in control, 20.0 µg/ml Cr(VI) and 20.0 µg/ml Mo(VI) exposed Oregon R⁺ larvae for 48 h (D). Extreme right panel represents overlayed images of H2 (green) and DAPI (blue) stained cells. Scale bar: 20 µm.</p

DEVDase (caspase 3-like) activity in the hemocytes of <i>Drosophila</i> larvae exposed to Cr(VI).

<p>Graphical representation of DEVDase activity (%) in the hemocytes of Oregon R⁺ larvae after their exposure to Cr(VI) (A). Data represent mean ± SD (n = 3) (50 larvae in each replicate). Significance in comparison to control was ascribed as *<i>p</i><0.05; **<i>p</i><0.01 and ***<i>p</i><0.001. Representative confocal images of hemocytes from control and 20.0 µg/ml Cr(VI) exposed Oregon R⁺ larvae for 48 h (B). Extreme right panel represents overlayed images of H2 (green), DAPI (blue) and cleaved caspase-3 (red) antibody stained cells. Scale bar: 20 µm.</p

Effect of NAC, L-NAME, SNP on THC and apoptosis in <i>Drosophila</i> hemocytes after Cr(VI) exposure.

<p>Graphical representation of total hemocyte number (%) in 20.0 µg/ml of Cr(VI) exposed Oregon R⁺ larvae along with 10 mg/ml N-acetylcysteine (NAC) or 100 mM N-nitro-L-arginine methyl ester (L-NAME) or 50 µM sodium nitroprusside (SNP) after 24 and 48 h (A1). Representative confocal images of hemocytes from control, 20.0 µg/ml Cr(VI), 20.0 µg/ml Cr(VI) with 10 mg/ml NAC, 20.0 µg/ml Cr(VI) with 100 mM L-NAME and 20.0 µg/ml Cr(VI) with 50 µM SNP exposed Oregon R⁺ larvae after 48 h (A2). Scale bar: 20 µm. Twenty larvae were taken for each replicate. Graphical representation of percent AV positive hemocytes in <i>Drosophila</i> larvae exposed to 20.0 µg/ml Cr(VI) along with 10 mg/ml NAC or 100 mM L-NAME or 50 µM SNP for 24 and 48 h respectively (B1). Dot plots showing Annexin V-FITC and PI staining in the hemocytes of control (a), 20.0 µg/ml Cr(VI) (b), 20.0 µg/ml Cr(VI) with 10 mg/ml NAC (c), 20.0 µg/ml Cr(VI) with 100 mM L-NAME (d) and 20.0 µg/ml Cr(VI) with 50 µM SNP (e) exposed Oregon R⁺ larvae after 48 h (B2). Fifty larvae were taken for each replicate. Data represent mean ± SD (n = 3). Significant differences were ascribed as **<i>p</i><0.01; ***<i>p</i><0.001 in comparison to control and <b>^#</b><i>p</i><0.05; <b>^##</b><i>p</i><0.01 and <b>^###</b><i>p</i><0.001 as compared to 20.0 µg/ml Cr(VI) exposure.</p

Generation of free radicals in the hemocytes of <i>Drosophila</i> larvae exposed to Cr(VI).

<p>Graphs showing O₂⁻ generation (A), H₂O₂ generation (B) and ONOO⁻ generation (C) in the hemocytes from control and Cr(VI) exposed Oregon R⁺ larvae. Values are mean ± SD (n = 3) (50 larvae in each replicate). Statistically significant differences were ascribed as *<i>p</i><0.05; **<i>p</i><0.01 and ***<i>p</i><0.001 in comparison to control.</p

Inhibited phagocytic activity of the hemocytes of Cr(VI) exposed <i>Drosophila</i> larvae.

<p>Graph showing inhibition of phagocytic activity (%) in the hemocytes of Cr(VI) exposed Oregon R⁺ larvae. Bar graphs represent mean ± SD (n = 3) (10 larvae in each replicate). Statistical significance was *<i>p</i><0.05; **<i>p</i><0.01 and ***<i>p</i><0.001 as compared to control.</p

Poor resistance of Cr(VI) exposed Oregon R⁺ larvae against <i>Ecc</i>15 infection.

<p>Survival (%) of <i>Drosophila</i> larvae that were exposed to Cr(VI) for 24 (A) and 48 (B) h followed by <i>Ecc</i>15 infection indicating resistance of an organism. Each survival curve in the graph represents mean survival of larvae from three independent experiments having 100 larvae in each and statistical significance was ascribed as **<i>p</i><0.01 and ***<i>p</i><0.001 as compared to control.</p

Schematic representation of Cr(VI)-induced alterations on cellular immunity in <i>D. melanogaster</i>.

<p>Cr(VI) altered cellular innate immune response through O₂^.−/ONOO<b>⁻</b> mediated oxidative stress in the hemocytes of exposed organism. The induction of oxidative stress leads to caspase-3 activation vis a vis apoptosis in the hemocytes which results into reduction in total hemocyte population in exposed organism. The altered immunity was manifested by decreased resistance of these organisms against pathogenic <i>Ecc</i>15 infection. Cr(VI) induced suppression of cellular immunity was subsequently modulated/rescued by over-expressing <i>sod</i> in <i>Drosophila</i> hemocytes.</p

Heat Shock Protein-70 (Hsp-70) Suppresses Paraquat-Induced Neurodegeneration by Inhibiting JNK and Caspase-3 Activation in <i>Drosophila</i> Model of Parkinson's Disease

<div><p>Parkinson's disease (PD) is one of the most common neurodegenerative disorders with limited clinical interventions. A number of epidemiological as well as case-control studies have revealed an association between pesticide exposure, especially of paraquat (PQ) and occurrence of PD. Hsp70, a molecular chaperone by function, has been shown as one of the modulators of neurological disorders. However, paucity of information regarding the protective role of Hsp70 on PQ-induced PD like symptoms led us to hypothesize that modulation of <i>hsp70</i> expression in the dopaminergic neurons would improve the health of these cells. We took advantage of <i>Drosophila</i>, which is a well-established model for neurological research and also possesses genetic tools for easy manipulation of gene expression with limited ethical concern. Over-expression of <i>hsp70</i> was found to reduce PQ-induced oxidative stress along with JNK and caspase-3 mediated dopaminergic neuronal cell death in exposed organism. Further, anti-apoptotic effect of <i>hsp70</i> was shown to confer better homeostasis in the dopaminergic neurons of PQ-exposed organism as evidenced by their improved locomotor performance and survival. The study has merit in the context of human concern since we observed protection of dopaminergic neurons in PQ-exposed organism by over-expressing a human homologue of <i>hsp70, HSPA1L,</i> in these cells. The effect was parallel to that observed with <i>Drosophila hsp70.</i> These findings reflect the potential therapeutic applicability of <i>hsp70</i> against PQ-induced PD like symptoms in an organism.</p></div

Correlation among ROS generation and % survival in PQ-exposed <i>Drosophila.</i>

<p>Negative correlation was observed between generation of ROS (O₂⁻/ONOO⁻) and percent survival in different strains of <i>Drosophila</i> exposed to 20 mM PQ for 24 h. n = 4 degree of freedom; <b>**</b><i>p</i><0.01.</p